Transient-state kinetic studies of the interactions of substrates and modified substrate derivatives with the enzymes enolase, creatine kinase, carboxypeptidase A, carbonic anhydrase and alkaline phosphatase are proposed. The purpose of this study is to identify reactive intermediates, the investigation of which should lead to clarification of some fundamental problems concerning enzyme catalysis. The study will employ principally temperature-jump relaxation, stopped-flow and combined flow-perturbation techniques. Analysis of the rate behavior of intermediates will allow us to describe the complex overall reactions in terms of sequences of elementary steps. Comparison of changes in the rate and thermodynamic parameters for the elementary steps with changes in overall reaction behavior of the system as various reagents are substituted will allow us to probe the reaction mechanism at the molecular level. The study is directed specifically towards elucidating structure-function relationships in metal ion activators and the mechanics of substrate recognition in enzyme catalysis.